Due to the ever-growing demand for healthy living, there is a keen interest in materials capable of killing harmful microorganisms. Such materials could be used to coat surfaces of common objects touched by people in everyday lives, e.g., door knobs, children toys, computer keyboards, telephones, etc., to render them antiseptic and thus unable to transmit bacterial infections. Since ordinary materials are not antimicrobial, their modification is required. For example, surfaces chemically modified with poly(ethylene glycol) and certain other synthetic polymers can repel (although not kill) microorganisms (Bridgett, M. J., et al, S. P. (1992) Biomaterials 13, 411–416. Arciola, C. R., et al Alvergna, P., Cenni, E. & Pizzoferrato, A. (1993) Biomaterials 14, 1161–1164. Park, K. D., Kim, Y. S., Han, D. K., Kim, Y. H., Lee, E. H. B., Suh, H. & Choi, K. S. (1998) Biomaterials 19, 851–859.)
Alternatively, materials can be impregnated with antimicrobial agents, such as antibiotics, quarternary ammonium compounds, silver ions, or iodine, that are gradually released into the surrounding solution over time and kill microorganisms there (Medlin, J. (1997) Environ. Health Persp. 105, 290–292; Nohr, R. S. & Macdonald, G. J. (1994) J. Biomater. Sci., Polymer Edn. 5, 607–619 Shearer, A. E. H., et al (2000) Biotechnol. Bioeng. 67, 141–146.). Although these strategies have been verified in aqueous solutions containing bacteria, they would not be expected to be effective against airborne bacteria in the absence of a liquid medium; this is especially true for release-based materials, which are also liable to become impotent when the leaching antibacterial agent is exhausted.
Infection is a frequent complication of many invasive surgical, therapeutic and diagnostic procedures. For procedures involving implantable medical devices, avoiding infection can be particularly problematic because bacteria can develop into biofilms, which protect the microbes from clearing by the subject's immune system. As these infections are difficult to treat with antibiotics, removal of the device is often necessitated, which is traumatic to the patient and increases the medical cost.
Any material left embedded in the body provides a surface for accumulation of infectious microorganisms, particularly bacteria and occasionally fungi. This is understood to take place through the formation of biofilms. A biofilm is a type of fouling that occurs when microorganisms attach to surfaces and secrete a hydrated polymeric matrix that surrounds them. Microorganisms existing in a biofilm, termed sessile, grow in a protected environment that insulates them from attack from antimicrobial agents. These sessile communities can give rise to nonsessile individuals, termed planktonic, which rapidly multiply and disperse. These planktonic organisms are responsible for invasive and disseminated infections. They are the targets of antimicrobial therapy. Conventional treatments fail to eradicate the sessile communities rooted in the biofilm. Biofilms are understood to be a frequently occurring reservoir for infectious agents. The biology of biofilms is described in more detail in “Bacterial biofilms: a common cause of persistent infection,” J. Costerson, P. Stewart, E. Greenberg, Science 284: 1318–1322 (1999), and “The riddle of biofilm resistance,” K. Lewis, Antimicrob. Agents Chemother., 45, 999–1007 (2001), incorporated herein by reference.
Biofilms develop preferentially on inert surfaces or on non-living tissue, and occur commonly on medical devices and devascularized or dead tissues. Biofilms have been identified on sequestra of dead bone and on bone grafts, from which they can incite an invasive infection called osteomyelitis that can kill even more bone. Biofilms have been also identified on living, hypovascular tissues such as native heart valves, where they are responsible for the devastating infection called endocarditis where the microorganism not only can colonize distant locations by seeding throughout the bloodstream, but also can destroy the heart valve itself. Infections involving implanted medical devices generally involve biofilms, where a sessile community provides a reservoir for an invasive infection. The presence of microorganisms in a biofilm on a medical device represents contamination of that foreign body. The elicitation by the biofilm of clinically perceptible host responses constitutes an infection.
The development of an infection from an area of contamination is consistent with the natural history of biofilm growth and development. Biofilms grow slowly, in one or more locations, colonized by one or a plurality of microorganisms. The pattern of biofilm development involves initial attachment of a microorganism to a solid surface, the formation of microcolonies attached to the surface, and finally the differentiation of the microcolonies into exopolysaccharide-encased mature biofilms. Planktonic cells are released from biofilms in a natural pattern of programmed detachment, so that the biofilm serves as a nidus for multiple, recurrent acute invasive infections. Antibiotics typically treat the infection caused by the planktonic organisms, but fail to kill those sessile organisms protected in the biofilm.
Sessile microorganisms also give rise to localized symptoms, releasing antigens and stimulating antibody production that activates the immune system to attack the biofilm and the area surrounding it. Antibodies and host immune defenses are ineffective in killing the organisms in the biofilm, even though these organisms have elicited the antibody and related immune response. The cytotoxic products of the host's immunologically activated cells can be directed towards the host's own tissues. This phenomenon is seen in the mouth, where the host's response to the dental biofilm can inflame tissues surrounding the teeth and give rise to periodontitis. This phenomenon can also give rise to local inflammation around implanted medical devices and bone resorption with loosening of orthopedic and dental implants.
While host defenses may hold invasive infections in check by controlling the proliferation of planktonic organisms, this favorable equilibrium presupposes an intact immune system. Many patients in a hospital setting have compromised immune systems, rendering them more vulnerable to invasive infections once a biofilm community has become established. Patients requiring implantable medical devices may likewise have compromised immune systems, whether on a short-term or long-term basis. A poorly functioning immune system puts the host at greater risk for initial formation of a contaminated biofilm around a medical device and for the invasion of planktonic organisms into the surrounding tissues and the system. Once the planktonic organisms mount a full-scale infection, the immunocompromised host will be less likely to contain and control it, with potentially lethal results.
Protected from antibiotic treatment and host defenses, the microorganisms in a biofilm typically cause recurrent infections and low-grade local symptoms. The biofilm, once established, can only be eradicated surgically. When a foreign object becomes contaminated with microorganisms, the only way to eliminate local and systemic infection may be to remove the contaminated foreign article. If the material being removed is essential for health, a similar article may need to be replaced in the same location; the replacement article will be especially prone to infection because of the residual microorganisms in the area.
Since the difficulties associated with eliminating biofilm-based infections are well-recognized, a number of technologies have developed to treat surfaces or fluids bathing surfaces to prevent or impair biofilm formation. Biofilms adversely affect medical systems and other systems essential to public health such as water supplies and food production facilities. A number of technologies have been proposed that treat surfaces with organic or inorganic materials to interfere with biofilm development. For example, various methods have been employed to coat the surfaces of medical devices with antibiotics (See e.g. U.S. Pat. Nos. 4,107,121, 4,442,133, 4,895,566, 4,917,686, 5,013,306, 4,952,419, 5,853,745 and 5,902,283) and other bacteriostatic compounds (See e.g U.S. Pat. Nos. 4,605,564, 4,886,505, 5,019,096, 5,295,979, 5,328,954, 5,681,575, 5,753,251, 5,770,255, and 5,877,243). Despite these technologies, contamination of medical devices and invasive infection therefrom continues to be a problem.
Infectious organisms are ubiquitous in the medical environment, despite vigorous efforts to maintain antisepsis. The presence of these organisms can result in infection of hospitalized patients and medical personnel. These infections, termed nosocomial, often involve organisms more virulent and more unusual than those encountered outside the hospital. In addition, hospital-acquired infections are more likely to involve organisms that have developed resistance to a number of antibiotics. Although cleansing and anti-bacterial regimens are routinely employed, infectious organisms readily colonize a variety of surfaces in the medical environment, especially those surfaces exposed to moisture or immersed in fluid. Even barrier materials, such as gloves, aprons and shields, can spread infection to the wearer or to others in the medical environment. Despite sterilization and cleansing, a variety of metallic and non-metallic materials in the medical environment can retain dangerous organisms trapped in a biofilm, thence to be passed on to other hosts.
Any agent used to impair biofilm formation in the medical environment must be safe to the user. Certain biocidal agents, in quantities sufficient to interfere with biofilms, also can damage host tissues. Antibiotics introduced into local tissue areas can induce the formation of resistant organisms which can then form biofilm communities whose planktonic microorganisms would likewise be resistant to the particular antibiotics. Any anti-biofilm or antifouling agent must furthermore not interfere with the salubrious characteristics of a medical device. Certain materials are selected to have a particular type of operator manipulability, softness, water-tightness, tensile strength or compressive durability, characteristics that cannot be altered by an agent added for anti-microbial effects.
As a further problem, it is possible that materials added to the surfaces of implantable devices to inhibit contamination and biofilm formation may be thrombogenic. Some implantable materials are of themselves thrombogenic. For example, it has been shown that contact with metal, glass, plastic or other similar surfaces can induce blood to clot. Heparin compounds, which are known to have anticoagulant effects, have therefore been applied to certain medical devices prior to implantation. However, there are few known products in the medical arsenal whose antimicrobial effects are combined with antithrombogenic effects. This combination would be particularly valuable to treat those medical devices that reside in the bloodstream, such as heart valves, artificial pumping devices (“artificial hearts” or left ventricular assist devices), vascular grafting prostheses and vascular stents. In these settings, clot formation can obstruct the flow of blood through the conduit and can furthermore break off pieces called emboli that are carried downstream, potentially blocking circulation to distant tissues or organs.
Biofilm formation has important public health implications. Drinking water systems are known to harbor biofilms, even though these environments often contain disinfectants. Any system providing an interface between a surface and a fluid has the potential for biofilm development. Water cooling towers for air conditioners are well-known to pose public health risks from biofilm formation, as episodic outbreaks of infections like Legionnaires' Disease attest. Turbulent fluid flow over the surface does not provide protection: biofilms can form in conduits where flowing water or other fluids pass, with the effects of altering flow characteristics and passing planktonic organisms downstream. Industrial fluid processing operations have experienced mechanical blockages, impedance of heat transfer processes, and biodeterioration of fluid-based industrial products, all attributable to biofilms. Biofilms have been identified in flow conduits like hemodialysis tubing, and in water distribution conduits. Biofilms have also been identified to cause biofouling in selected municipal water storage tanks, private wells and drip irrigation systems, unaffected by treatments with up to 200 ppm chlorine.
Biofilms are a constant problem in food processing environments. Food processing involves fluids, solid material and their combination. As an example, milk processing facilities provide fluid conduits and areas of fluid residence on surfaces. Cleansing milking and milk processing equipment presently utilizes interactions of mechanical, thermal and chemical processes in an air-injected clean-in-place methods. Additionally, the milk product itself is treated with pasteurization. In cheese producing, biofilms can lead to the production of calcium lactate crystals in Cheddar cheese. Meat processing and packing facilities are in like manner susceptible to biofilm formation. Non-metallic and metallic surfaces can be affected. Biofilms in meat processing facilities have been detected on rubber “fingers,” plastic curtains, conveyor belt material, evisceration equipment and stainless steel surfaces. Controlling biofilms and microorganism contamination in food processing is hampered by the additional need that the agent used not affect the taste, texture or aesthetics of the product.
There exists, therefore, a need to be able to render general surfaces bactericidal. General surface coating/derivatization procedures have been developed that should be applicable to most materials regardless of their nature.